Key Laboratory of Organ Regeneration and Reconstruction, State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China.
Institute for Stem Cell and Regeneration, Chinese Academy of Sciences, Beijing, China.
Cell Prolif. 2024 Jun;57(6):e13599. doi: 10.1111/cpr.13599. Epub 2024 Jan 12.
The embryo-like structures (embryoids) constructed by aggregating embryonic stem cells (ESCs) and trophoblast stem cells (TSCs) have provided revolutionary tools for studying the intricate interaction between embryonic and extra-embryonic tissues during early embryonic development, which has been achieved in mice. However, due to the opposite dependence on some signalling pathways for in vitro culture of human ESCs (hESCs) and TSCs (hTSCs), particularly WNT and TGFβ signalling pathways, which limits the construction of human post-implantation embryoids by aggregating hESCs and hTSCs. To overcome this challenge, here, by screening 1639 chemicals, we found that an inhibitor of integrated stress response, ISRIB, can replace WNT agonists and TGFβ inhibitors to maintain the stemness and differentiation capacity of hTSCs. Thus, we developed an ISRIB-dependent in vitro culture medium for hTSCs, namely nTSM. Furthermore, we demonstrated that ISRIB could also maintain the hESC stemness. Using a 3D co-culture system (hESCs and hTSCs aggregate, ETA), we demonstrated that a 1:1 mixture of hESC culture medium (ESM) and nTSM improved the cell proliferation and organisation of both hESC- and hTSC-compartments and the lumenogenesis of hESC-compartment in ETAs. Overall, our study provided an ISRIB-dependent system for co-culturing hESCs and hTSCs, which facilitated the construction of human embryoids by aggregating hESCs and hTSCs.
由聚集胚胎干细胞（ESCs）和滋养层干细胞（TSCs）构建的类胚胎结构（胚状体）为研究早期胚胎发育中胚胎和胚胎外组织之间复杂的相互作用提供了革命性的工具，这在小鼠中已经实现。然而，由于人类胚胎干细胞（hESCs）和滋养层干细胞（hTSCs）的体外培养对某些信号通路存在相反的依赖性，特别是 WNT 和 TGFβ 信号通路，这限制了通过聚集 hESCs 和 hTSCs 构建人类植入后胚状体。为了克服这一挑战，在这里，我们通过筛选 1639 种化学物质，发现一种整合应激反应抑制剂 ISRIB 可以替代 WNT 激动剂和 TGFβ 抑制剂来维持 hTSCs 的干性和分化能力。因此，我们开发了一种依赖于 ISRIB 的 hTSCs 体外培养培养基，即 nTSM。此外，我们证明 ISRIB 还可以维持 hESC 的干性。使用 3D 共培养系统（hESC 和 hTSC 聚集，ETA），我们证明了 hESC 培养基（ESM）和 nTSM 的 1:1 混合物可提高 ETAs 中 hESC 和 hTSC 区室的细胞增殖和组织以及 hESC 区室的管腔形成。总体而言，我们的研究提供了一种依赖于 ISRIB 的 hESC 和 hTSC 共培养系统，促进了通过聚集 hESC 和 hTSC 构建人类胚状体。
